Yin Cui-Ping, Guan Shang-Hui, Zhang Bo, Wang Xue-Xin, Yue Shou-Wei
Department of Physical Medicine and Rehabilitation, Qilu Hospital of Shandong University, Jinan, Shandong 250012, P.R. China.
Department of Radiation Oncology, Qilu Hospital of Shandong University, Jinan, Shandong 250012, P.R. China.
Mol Med Rep. 2015 Nov;12(5):7123-31. doi: 10.3892/mmr.2015.4267. Epub 2015 Aug 28.
Hypoxic conditions regulate several metabolic enzymes and transcription factors that are involved in cancer, ischemia and pulmonary diseases. The Ras homolog (Rho) family, including Rho member A (RhoA), is involved in reorganization of the actin cytoskeleton, cell migration and in the regulation of apoptosis and gene transcription. The aim of the present study was to investigate the expression of hypoxia‑inducible factor (HIF)‑α and the activity of RhoA in PC12 neuroblastoma cells under hypoxic conditions. The upregulation of HIF‑α and RhoA by hypoxia was determined using reverse transcription‑quantitative polymerase chain reaction and western blot assays, cell apoptosis was analyzed using flow cytometry, and the activity of caspase 3 was examined using a western blot assay and caspase 3 activity assay kit. The PC12 cells were induced to apoptosis following exposure to hypoxia, and exhibited increased expression of HIF‑α and increased mRNA and protein expression levels of RhoA. The overexpression of HIF‑α attenuated the hypoxia‑induced apoptosis of the PC12 cells. In addition, RhoA knockdown using small interfering RNA abrogated the antagonism of HIF‑1α towards hypoxia‑induced apoptosis. The results of the present study confirmed the protective role of HIF‑1α and RhoA in hypoxia‑induced PC12 cell apoptosis, and that the upregulation of HIF‑1α by hypoxia is RhoA‑dependent.
缺氧条件可调节多种参与癌症、缺血和肺部疾病的代谢酶和转录因子。Ras同源物（Rho）家族，包括Rho成员A（RhoA），参与肌动蛋白细胞骨架的重组、细胞迁移以及细胞凋亡和基因转录的调节。本研究的目的是探讨缺氧条件下PC12神经母细胞瘤细胞中缺氧诱导因子（HIF）-α的表达及RhoA的活性。采用逆转录-定量聚合酶链反应和蛋白质印迹法检测缺氧对HIF-α和RhoA的上调作用，采用流式细胞术分析细胞凋亡，采用蛋白质印迹法和半胱天冬酶3活性检测试剂盒检测半胱天冬酶3的活性。PC12细胞在缺氧后被诱导凋亡，并表现出HIF-α表达增加以及RhoA的mRNA和蛋白质表达水平升高。HIF-α的过表达减弱了缺氧诱导的PC12细胞凋亡。此外，使用小干扰RNA敲低RhoA可消除HIF-1α对缺氧诱导凋亡的拮抗作用。本研究结果证实了HIF-1α和RhoA在缺氧诱导的PC12细胞凋亡中的保护作用，且缺氧引起的HIF-1α上调依赖于RhoA。
